Embedded Files
Anna Burns
Y109C TAT --> TGT
T1121C ACA --> TGT
Y109C
Y109C
Got through TENS Prep and Ethanol precipitation
NanoDrop of MidiPrep Product - 06/13/2022
NanoDrop of MidiPrep Product - 06/13/2022
Y109C_CFTR_pGHE_1 : 8905.1 ng/uL
Y109C_CFTR_pGHE_2 : 14895.6 ng/uL
Sequencing - 06/29/2022
Sequencing - 06/29/2022
Results: Successfully mutated TAT to TGT
T1121C
T1121C
PCR Mutagenesis - 07/21/2022
PCR Mutagenesis - 07/21/2022
Template DNA was midiprep product
Y109C_CFTR_pGHE_1 : 8905.1 ng/uL Diluted both to 25ng/uL
Y109C_CFTR_pGHE_2 : 14895.6 ng/uL
Gel Electrophoresis - 07/23/2022
Gel Electrophoresis - 07/23/2022
Gel Analysis PCR was successful, did Dpn1 digestion and will be proceeding with transformation
Gel Analysis PCR was successful, did Dpn1 digestion and will be proceeding with transformation
Circularization and Nanodropping - 07/25/2022
Circularization and Nanodropping - 07/25/2022
Nanodrop concentration:
Y109C/T1121C - 1 : 665.87 ng/uL Diluted to 40ng/uL for transcription
Y109C/T1121C - 2 : 666.17 ng/uL
Transformation - 07/26/2022
Transformation - 07/26/2022
There was some extra liquid in Y109C/T1121C-1, so I'm afraid I accidentally doubled the amount of JM109 cells in this tube and accidentally skipped another tube. We will see though.
Finished at 9:15 pm
Y109C/T1121-1
Y109C/T1121-1
Scattered colony growth, there are several usable ones though.
Y109C/T1121C -2
Y109C/T1121C -2
Overconcentrated bacterial growth, something failed. I will be redoing transformation at a later date.
Negative Control
Negative Control
No bacterial growth, indicating a sterile procedure
Redoing Transformation - 08/15/2022
Redoing Transformation - 08/15/2022
Y109C/T1121C - 2 Used 30uL JM109
Very nice bacterial growth.
Single Colony Inoculation - 07/28/2022
Single Colony Inoculation - 07/28/2022
Placed in the incubator at 6:51 pm, will remove between 9:51 and 10:51 am
Y109C/T1121C - 1
Y109C/T1121C - 1
Cloudy broth indicating the bacterial culture was present and single colony inoculation was successful
Negative Control
Negative Control
No bacterial growth, indicating a sterile procedure.
Single Colony Inoculation - 08/16/2022
Single Colony Inoculation - 08/16/2022
Placed in the incubator at 6:48 pm, will remove around 9:48am - 10:48am
Y109C/T1121C - 2
Y109C/T1121C - 2
Cloudy broth shows adequate bacterial growth
Negative Control
Negative Control
Clear broth means no bacterial growth, indicating a sterile procedure.
Maxiprep - 07/29/2022
Maxiprep - 07/29/2022
Removed from the incubator at 10:29 am.
The pellets were too small for my taste, so I added 1/10 total volume of sodium acetate buffer and let it sit overnight.
Resuspended in 1mL MBG H2O on
Y109C/T1121C - 1
Y109C/T1121C - 1
First pellet
Y109C/T1121C - 2
Y109C/T1121C - 2
First Pellet
Y109/T1121C - 2
Y109/T1121C - 2
Final pellet
Nanodrop - 08/09/2022 , 08/24/2022
Nanodrop - 08/09/2022 , 08/24/2022
Y109C/T1121C - 1: 2174.6 ng/uL
Y109C/T1121C - 2: 3653.47ng/uL
Gel Electrophoresis - 08/10/2022
Gel Electrophoresis - 08/10/2022
Gel Analysis Lane 9
Some RNA contamination, I will add some more RNAse
Sequencing - 08/11/2022 , 08/24/2022
Sequencing - 08/11/2022 , 08/24/2022
Failed, see doc for more details.
Did not run a gel first, nanodrop showed the presence of dsDNA.
Page updated
Report abuse